Discovery of Genes Expressed in Basal Endosperm Transfer Cells in Maize Using 454 Transcriptome Sequencing

Endosperm in a developing seed of maize is a major storage tissue for starch and proteins, and is of immense economic significance for its role in food, feed, and biofuel production. Of the total four cell types in developing endosperm, basal endosperm transfer cells (BETCs) are known to perform a major role in solute acquisition from post-phloem maternal tissues of developing seeds of the plant. To identify all transcripts expressed in the BETCs, we employed here a simplified microdissection method to isolate the BETCs from developing kernels at 12 days after pollination. Total RNA was extracted from such BETCs and used for cDNA synthesis. The 5,790 reads of cDNAs with an average length of 218 nucleotides were generated through 454 sequencing of a partial (1/16) run of a picotiter plate. After trimming, a total of 4,505 reads were used for clustering and assembly that led to 682 contigs and 1,791 singletons. A select set of genes showing high abundance in the above transcript reads were tested and validated for the BETC-specific expression by qPCR and by in situ hybridization. Positive results from these tests and the undetectability of expression of the marker genes relating to the embryo surrounding region located adjacent to the BETCs suggested that our microdissection method was successful in harvesting the BETC-enriched cells. Functional annotation and categorization of the 2,473 unique sequences showed high abundance of transcripts related to mitochondrial activity, stress stimulus, alkaloid biosynthesis related to defense genes and various signaling functions essential for seed development.